Objective/Rationale:
A growing number of epidemiological and experimental animal studies implicate neuroinflammatory mechanisms in the progressive death of midbrain dopamine (DA) neurons in Parkinson’s Disease. Our recent work indicates that 50% of the DA neurons destined to die after a neurotoxic or an inflammatory insult can be rescued using engineered dominant negative human Tumor Necrosis Factor peptides (DN-TNFs) which selectively block the activity of the soluble form of the pro-inflammatory cytokine TNF. These preclinical studies validate TNF as a potential target for development of new therapeutic approaches to slow or prevent the progressive degeneration of the nigrostriatal pathway. However, since protein-based inhibitors have relatively short half-lives and do not cross the blood-brain barrier, they must be delivered directly into the midbrain via a chronically implanted device. To circumvent the inherent limitations associated with this drug delivery approach, we now propose to develop constitutive and inducible lentiviral DN-TNF vectors as a novel anti-inflammatory gene therapy approach for neuroprotection of nigral DA neurons.
Project Description:
Constitutive and inducible lentiviral vectors encoding human DN-TNF sequences will be developed and validated. Cell-type specific expression of hDN-TNFs achieved after infection of DA neuron cell lines or mixed primary neuron/glia cultures and the ability of hDN-TNF-expressing cells to defend against neurotoxic insults in culture will be investigated using various cell biological and imaging assays. The extent to which lenti-hDN-TNFs can block lipid peroxidation, mitochondrial dysfunction and the progressive loss of DA neurons in a pre-clinical model of PD will be established. Tetracycline-inducible lenti-DN-TNFs will be used to investigate the length of time of TNF inhibition required to achieve lasting neuroprotection and whether delayed TNF inhibition can still neuroprotect in vivo.
Relevance to Diagnosis/Treatment of Parkinson’s Disease:
A potential new therapeutic modality (in vivo anti-TNF gene therapy) for neuroprotection of the nigrostriatal pathway may emerge from this work that could be advanced to clinical trials expediently.
Anticipated Outcome:
These studies will enable us to determine the critical stage and molecular pathways by which TNF contributes to DA neuron dysfunction and death. Completion of these studies will provide both research tools and conclusive answers regarding the feasibility and efficacy of anti-TNF gene therapy in attenuating progressive nigrostriatal pathway degeneration.